Biological Effects And Mechanism Of Sulforaphane On Colorectal Cancer

ObjectiveMalignant tumors are increasingly becoming an important factor threatening the survival of all mankind,among which colorectal cancer ranks top in both morbidity and mortality worldwide.Colorectal cancer ranks third in the world in new cancers with a 10.2 percent incidence,according to the 2018 global oncology annual report.Worldwide,the case fatality rate of 9.2% of colorectal cancer patients is second only to that of lung cancer(18.4%).Colorectal cancer is the third leading cause of morbidity and the fourth leading cause of mortality in men.Colorectal cancer is the second leading cause of morbidity and mortality in women.These data fully show that colorectal cancer is a growing threat to human health.Therefore,it is increasingly important to study the mechanism of colorectal cancer occurrence and development and to find new strategies for colorectal cancer treatment and prevention.In recent years,with the in-depth research on the mechanism of tumorigenesis and development,it has been confirmed that dietary habits have a significant impact on the morbidity and mortality of certain tumors,especially those of the digestive system.At the same time,a reasonable diet for the prevention of cancer is also of great help.Epidemiological studies have shown that increasing the intake of cruciferous vegetables can effectively reduce the incidence of a variety of tumors.Sulforaphane is a chemical active ingredient in cruciferous vegetables,especially broccoli and cauliflower.Sulforaphane has been found to be effective in inhibiting the development of several types of cancer,including lung,breast,prostate,colorectal,kidney and esophageal cancers.Sulforaphane can exert tumor cytotoxicity through various mechanisms,such as inducing phase II enzymes that play a key role in the elimination of tumor mutagens,activating apoptosis and inducing cell cycle arrest.The main task of this project is to study the effect of sulforaphane on the biological behavior of colorectal cancer cells,and to explore the mechanism of these phenomena,so as to provide new strategies and ideas for the prevention of colorectal cancer.MethodsIn this study,HCT116 and SW480 cell lines were mainly used for research.The study first explored the dose-time relationship between sulforaphane and cell apoptosis and cell activity in colorectal cancer cell lines.Sulforaphane at 0?M,10?M,20?M and 30?M was designed to treat HCT116 and SW480,and the apoptosis ratio was detected by flow cytometry with annexin-v and PI double staining at 24 h,36h and 48 h time gradients.The effect of sulforaphane on proliferation and cell activity of HCT116 and SW480 cells was also detected by CCK-8 assay.Next,the causes of cell apoptosis and cell activity inhibition induced by sulforaphane were investigated.After treatment with sulforaphane,intracellular ROS levels were detected by flow cytometry,and the biological effects of sulforaphane on cells were studied by adding ROS scavenger(N-acetyl-L-cysteine,NAC).Next,the mechanism of sulforaphane-induced increase of ROS was investigated.In cell metabolism,endogenous ROS mainly come from electron transport chain,fat oxidation,NADPH oxidation,xanthine/hypoxanthine oxidation,nitric oxide synthesis,etc.Therefore,we add the above key metabolic enzyme inhibitors,including electron transport chain inhibitors(Rotenone,Rot),lipoxygenase inhibitors(Nordihydroguaiaretic acid,NDGA),NADPH oxidase inhibitors(Diphenylenediodonium chloride,DPI),xanthine/ xanthine oxidation inhibitors(Allopurinol,Allo)and nitric oxide synthase inhibitors(1400 w.2 HCL,1400 w).Tumor cells were treated with sulforaphane and different inhibitors to identify pathways that may be involved in sulforaphane-induced endogenous ROS.After treatment with different inhibitors and sulforaphane,the levels of ROS and apoptosis were measured by flow cytometry.Finally,the effect of sulforaphane on cell signaling pathway was determined by western-blot.Results1.Sulforaphane can inhibit the cell proliferation and cell activity of colorectal cancer cell lines HCT116 and SW480 and induce apoptosis both in a time and dose dependent manner.2.Sulforaphane-induced apoptosis of colorectal cancer cell lines HCT116 and SW480 mainly by increasing intracellular ROS level.Scavenging ROS can effectively eliminate the effects of sulforaphane on apoptosis,cell proliferation and cell viability.3.The increase of endogenous ROS in the colorectal cancer cell lines HCT116 and SW480 after sulforaphane treatment was mainly through enhancing the activity of NADPH oxidase and xanthine oxidase.Inhibition of NADPH oxidase and xanthine oxidase activity can reduce the effect of sulforaphane on cell biology.4.Sulforaphane treatment of colorectal cancer cell lines HCT116 and SW480 activated the phosphorylation pathway PI3 K / AKT / mTOR.ConclusionAs a chemically active substance in vegetables such as broccoli,sulforaphane inhibits tumor cell proliferation and cell activity,and induces cell apoptosis.Mainly sulforaphane enhances intracellular NADPH oxidase and xanthine oxidase activity,produces a large amount of ROS,induces cell damage and apoptosis,and inhibits cell activity and cell proliferation.At the same time,sulforaphane activates the PI3K/AKT/mTOR phosphorylation pathway associated with cell survival.